Hepatitis C is an RNA virus containing a single-stranded positive-sense RNA genome of about 9,600 nts. The genes for the viral structural and non-structural proteins are flanked by 5′ and 3′ untranslated regions (UTRs), which are essential for genome replication. For example, the 5′ UTR contains an internal ribosome entry site (IRES) which is indispensable for the initiation of HCV polyprotein translation. HCV has been classified into six major genotypes, each comprising further subgroups, which differ in their sequence homology by more than 30%. The distribution of these genotypes differs geographically. For example, genotypes 1a and 1b are the most prevalent genotypes found within the U.S., while genotypes 2 and 3 are more prevalent in other countries.
Because of the sequence variability of HCV, the development of vaccines and therapeutic drugs, including RNAi-based therapeutics, that would be active against the majority of viruses, must take advantage of the rare conserved epitopes and sequences found among the viral genotypes and quasispecies. In fact, the mutability of HCV is such that even within an infected individual, the HCV virus exists as a swarm of variants or “quasispecies” of a predominant type rather than as a single entity.
Thus, to apply a gene-silencing-based strategy to the treatment or prevention of HCV infection, it is necessary to identify sufficiently conserved stretches of nucleotide sequence in this highly mutable virus. That is, since RNA interference is a sequence-specific effect, therapeutic or prophylactic RNAi molecules must be specific for HCV target sequences, despite the fact that hepatitis C viral genomes are highly variable. While HCV target sequence conservation is an important consideration in the design of sequence-specific anti-HCV prophylactic or therapeutic modalities such as RNAi or antisense, e.g., some of the highly conserved regions of the HCV genome such as the 5′ UTR are known to be highly structured, while some regions of the viral genome are present in the infected cell in association with proteins which make them largely inaccessible to antisense or RNAi. The lack of a readily available HCV animal model and problems with various HCV cell culture models, e.g., the absence or deficiencies in viral infection or replication models, have hindered the development of anti-HCV pharmaceuticals of all types.
Despite well over a decade of research efforts, there are no vaccines available for HCV. As a consequence, the rate of new HCV infections around the world is extremely high. The WHO estimates that globally 170 million individuals carry chronic HCV infections and that new infections are established at a rate of 3 to 4 million annually.
Chronic HCV infection induces liver inflammation, causing progressive liver disease that can lead to cirrhosis and hepatocellular carcinoma (liver cancer). Chronic HCV infection becomes established in 75%-85% of individuals experiencing an initial infection, and HCV-related liver failure is the most common indication cited for liver transplantation in the U.S. Chronic HCV infection in its early stages may cause only mild non-specific symptoms, such as fatigue, or be completely asymptomatic, leaving many infected individuals unaware that they carry a dangerous chronic infection.
Current therapies for HCV infection, which may include a 6 to 12 month regimen of pegylated interferon and ribavirin, can lead to a cure in a minority of patients. Response rates vary by HCV genotype, with genotype 2 and 3 patients exhibiting a 76% response rate to the current standard therapy while patients infected with genotype 1a and 1b having only a 46% response rate. Unfortunately, genotype 1 accounts for 60% of global infections and is the dominant strain in the U.S., Japan, and Western Europe. Complicating genotype 1 resistance to ribavarin and interferon is the fact that both drugs have side effect profiles that can require dose reduction or discontinuance of therapy when patients experience side effects. Further complicating patient outcomes is the fact that patients who fail an initial treatment regimen rarely respond favorably to a subsequent round of treatment with interferon and ribavarin.
Clinicians who treat HCV patients are hopeful that current and future research programs will yield options that improve the response rate for genotype 1 patients, which is currently less than 50% using ribavarin and interferon. New treatment options that have a more tolerable side effect profile would improve patient compliance and enable more patients to complete a full course of therapeutic intervention.
There remains a need for treatment options for HCV-exposed or infected patients, including for highly conserved nucleic acid-based molecules, including double-stranded RNAs and constructs encoding dsRNAs, capable of inhibiting the replication of HCV in mammalian cells. Such nucleic acid based anti-HCV therapeutic agents have the potential to improve patient response rates to therapy, improve adverse event profile, and eliminate or significantly delay the development of drug resistant escape mutant virus.